Severe tissue injury generates central sensitization (increased responsiveness of CNS nociceptive neurons to normal or sub-threshold afferent input) that contributes to hyperalgesia. Latent sensitization (LS) is a silent form of central sensitization that persists after tissue has healed and overt signs of hyperalgesia have resolved. LS can be revealed with opioid receptor antagonists or inverse agonists that rekindle or reinstate hyperalgesia. Thus, pain remission during LS is likely maintained by tonic opioid receptor activity that masks the pronociceptive components of LS. LS is important because it primes nociceptive systems such that, when inhibitory systems fail, a pain episode ensues. A key first step in understanding LS is to demonstrate the translational significance, and we now show that the opioid receptor inverse agonist, naloxone, can reinstate experimental pain when delivered 1 wk after the resolution of secondary hyperalgesia following first degree thermal injury. Specific Aim 1 tests the hypothesis that burn or surgery triggers LS and long-term opioid analgesia in humans. To further study the neurobiological mechanisms of LS, we will also use a mouse model that is long-lasting, powerful, broad range, repeatable, and translates to human studies. We found that mu opioid receptor (MOR) inverse agonists reinstated behavioral and molecular signs of hyperalgesia, even when administered months after tissue injury, and this required NMDA receptor activation of adenylyl cyclase type 1 (AC1). Our results are important because they suggest that any event, such as stress, that interferes with MOR analgesia during LS will lead to relapse of hyperalgesia in chronic pain syndromes in humans. Specific Aim 2 tests the hypothesis that MOR constitutive activity (MORCA) and/or activation of MOR, delta (DOR), or kappa (KOR) receptors by opioid peptides in the DH or rostroventromedial medulla maintains endogenous analgesia and thereby restricts LS to a state of pain remission. Specific Aim 3 determines the extent to which MORs inhibit spatially coordinated neural activity in the DH (using an innovative 64-channel field recording system) and synaptic strength in presynaptic terminals of primary afferent nociceptors or on DH neurons (using patch clamp electrophysiology) during LS. Specific Aim 4 then tests whether MORs specifically inhibit spinal NMDA receptor subunits (GluN2A or GluN2B) and/or Epac1 (exchange protein directly activated by cAMP, recently found to contribute to peripheral pain senstization) to block pain during LS. Completion of this project will bring us closer to our long-term goal of alleviating chronic pain b either: a) facilitating endogenous opioid analgesia, thus restricting LS within a state of remission; or b) extinguishing LS altogether, for example with a selective AC1 or Epac1 inhibitor. Our general model and hypothesis shares similarities with the concept of allostasis: a pathologically-elevated balance between opposing processes (MOR and LS) that facilitate each other by mutual feedback. Our long-term vision is a new conceptual strategy for chronic pain therapy, to restore homeostasis, where there is neither central sensitization nor MOR compensatory responses.